The main goal of the present project is the functional characterization of the NIT2 transcription factor (described as a regulator of the nitrogen metabolism in Neurospora crassa) as a putative regulator of the glycogen metabolism in this fungus. Previous results in our laboratory showed that in normal growth conditions, N. crassa accumulates glycogen in the late of exponential phase and decreases it in the early stationary phase. In a stressful situation, such as heat shock (from 30ºC to 45ºC) it is known that the fungus degrades glycogen. Moreover, in the same environmental condition the gsn transcript that codifies glycogen synthase enzyme (regulatory enzyme of biosynthesis) is severely reduced and recovered when the wild-type strain returns to the normal temperature (30ºC). Recently, we reported that the nit2KO strain presents a profile different from that described for the wild-type strain: a hiper-accumulation of glycogen was observed both before and after heat shock. In an attempt to understand the molecular mechanism involved in such regulation, the 5'-flanking regions of the gsn and gpn (codifies glycogen phosphorylase enzyme) genes were analyzed in relation to the presence of regulatory DNA elements for the NIT2 transcription factor. One motif was identified in the 5'-gsn and five motifs were identified in the 5'-gpn. In a preliminary analysis, it was observed that proteins present in crude cellular extract from wild-type mycelium grown at physiological conditions were able to bind to a DNA fragment from the gsn promoter containing the NIT2 consensus site. This project has as main objective to characterize the role of this transcription factor in regulating the glycogen metabolism.
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